Colorectal cancer early warning serum index and detection method and application thereof
An early warning technology for colorectal cancer, applied in the field of medical biology, can solve the problems of lack of serum indicators for early warning of colorectal cancer, and achieve the effect of improving sensitivity and specificity, high sensitivity, and strong stability
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example 1
[0050] Example 1. An ELISA kit for detecting the concentration of angiogenin (using a marker for detection), the components of the kit include:
[0051] 1) The coating buffer is 0.05M carbonate buffer, and its composition is 1.59g / L Na 2 CO 3 and 2.93g / LNaHCO 3 , the balance is deionized water; pH value is 8.5;
[0052] 2) Washing solution I is PBS buffer, its components are 140mM NaCl, 2.7mM KCl, 10mM Na 2 HPO 4 , 1.8mMKH 2 PO 4 , the balance is deionized water; pH value is 7.4;
[0053] 3) Washing solution II is PBS containing 1% (vol%) Tween-20, called PBST, and its components are PBS and 1% Tween-20; that is, add 1% (vol%) Tween in PBS to PBS -20;
[0054] 4) The blocking solution is PBS containing 1% (mass %) BSA, and its components are PBS and 1% BSA; that is, 1 g of BSA per 100 ml of PBS;
[0055] 5) The composition of the loading buffer is 50mM Tris-HCl, 0.5M KCl, 1% (mass %) BSA, 0.05% (volume %) Tween-20, and the balance is deionized water; the pH value is 7...
Embodiment 1
[0062] Embodiment 1, human angiopoietin detection kit detects the content of angiopoietin in the serum of healthy people
[0063] Adopt the human angiogenin detection kit described in example 1 to detect the content of angiogenin in the serum of healthy people; the specific detection method is as follows:
[0064] 1) Add mouse anti-human angiopoietin monoclonal antibody diluted 100 times in coating buffer to a 96-well ELISA plate, 100 μL per well, and coat at 4°C for 24 hours;
[0065] 2) Discard the liquid in the well, and wash three times with Washing Solution I for 5 minutes each time;
[0066] 3) Wash three times with washing solution II, 5 minutes each time;
[0067] 4) Add 200 μL of blocking solution to each well and block for 2 hours at room temperature;
[0068] 5) Repeat the washing of steps 2) and 3); that is, discard the liquid in the well; wash with washing solution I and washing solution II;
[0069] 6) Prepare the sample:
[0070] Prepare standard samples of ...
Embodiment 2
[0082] Example 2. Specificity analysis of angiogenin detection kit
[0083] The human angiopoietin detection kit described in Example 1 above was used to detect the content of other members of this family, including RNase 1, RNase 4, RNase 6, RNase 7 and RNase 8.
[0084] Compared with steps 1) to 16) of Example 1, the only difference is that the samples prepared in step 6) are 1000 pg / ml of human angiogenin, ribonuclease 1, ribonuclease 4, ribonuclease 6, RNase 7 and RNase 8; step 15 and step 16 are omitted); the rest are the same.
[0085] That is, the specific detection method is as follows: first, take a 96-well ELISA plate coated with angiopoietin monoclonal antibody, and discard the liquid in the well; wash the ELISA plate with washing solution I and washing solution II; then, prepare blood vessels Genitin (R&D Systems, product number: 265-AN), RNase 1 (Sangon Biotechnology Co., Ltd., product number: D626068), RNase 4 (Sangon Biotechnology Co., Ltd., product number: D62...
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